^{Skeletal Myoblast-Seeded Vascularized Tissue Scaffolds in the Treatment of a Large Volumetric Muscle Defect in the Rat Biceps Femoris Muscle}

Abstract

High velocity impact injuries can often result in loss of large skeletal muscle mass, creating defects devoid of matrix, cells, and vasculature. Functional regeneration within these regions of large volumetric muscle loss (VML) continues to be a significant clinical challenge. Large cell-seeded, space-filling tissue-engineered constructs that may augment regeneration require adequate vascularization to maintain cell viability. However, the long-term effect of improved vascularization and the effect of addition of myoblasts to vascularized constructs have not been determined in large VMLs. Here, our objective was to create a new VML model, consisting of a full-thickness, single muscle defect, in the rat biceps femoris muscle, and evaluate the ability of myoblast-seeded vascularized collagen hydrogel constructs to augment VML regeneration. Adipose-derived microvessels were cultured with or without myoblasts to form vascular networks within collagen constructs. In the animal model, the VML injury was created in the left hind limb, and treated with the harvested autograft itself, constructs with microvessel fragments (MVF) only, constructs with microvessels and myoblasts (MVF+Myoblasts), or left empty. We evaluated the formation of vascular networks in vitro by light microscopy, and the capacity of vascularized constructs to augment early revascularization and muscle regeneration in the VML using perfusion angiography and creatine kinase activity, respectively. Myoblasts (Pax7+) were able to differentiate into myotubes (sarcomeric myosin MF20+) in vitro. The MVF+Myoblast group showed longer and more branched microvascular networks than the MVF group in vitro, but showed similar overall defect site vascular volumes at 2 weeks postimplantation by microcomputed tomography angiography. However, a larger number of small-diameter vessels were observed in the vascularized construct-treated groups. Yet, both vascularized implant groups showed primarily fibrotic tissue with adipose infiltration, poor maintenance of tissue volume within the VML, and little muscle regeneration. These data suggest that while vascularization may play an important supportive role, other factors besides adequate vascularity may determine the fate of regenerating volumetric muscle defects.

Effects of BMP‐2 dose and delivery of microvascular fragments on healing of bone defects with concomitant volumetric muscle loss

“…MVF have been shown to improve tissue vascularization and integration; however, this has been primarily demonstrated in subcutaneous implantation, 20 skin flaps, 39 and dermal substitutes, 22,23 all of which are considerably thinner and thus closer to native vascular sources than the critical size bone defect used here. MVF have also been shown to increase tissue perfusion in VML models when delivered in collagen gel, 21,40 which allows for sprouting angiogenesis. 17,21,41,42 In this study, while in vitro viability of MVF was maintained in collagen sponge, the multicellular MVF first dissociated to single cells and then reformed networks.…”

“…MVF have also been shown to increase tissue perfusion in VML models when delivered in collagen gel, 21,40 which allows for sprouting angiogenesis. 17,21,41,42 In this study, while in vitro viability of MVF was maintained in collagen sponge, the multicellular MVF first dissociated to single cells and then reformed networks. This is not consistent with the sprouting angiogenesis typically observed within a collagen gel and may have contributed to the lack of effect of MVF on bone healing.…”

Skeletal Muscle Tissue Engineering: Biomaterials-Based Strategies for the Treatment of Volumetric Muscle Loss

“…Several researchers developed pre-vascularized TEMGs and evaluated their ability to participate in skeletal muscle regeneration in vivo [246,247,[264][265][266][267]. Li et al created TEMGs with collagen hydrogels seeded with adipose-derived microvessels with or without myoblasts [264]. Upon implantation into a full thickness biceps femoris defect, vascularized constructs were unable to prevent fibrosis and did not support muscle regeneration; this may suggest the supportive role that vascularization plays in muscle regeneration.…”

Muscle Stem Cell Niche Dysregulation in Volumetric Muscle Loss Injury

“…This is clearly a necessary criterion for creating a tissue engineered construct for skeletal muscle, as it is a highly vascularized tissue. There have been several studies which have shown tissue engineered strategies which promote successful formation of vessels after VML injury (M.-T. Li et al, 2017;VanDusen, Syverud, Williams, Lee, & Larkin, 2014). Interestingly, in our critically sized VML defect with no treatment we saw increased vascular volume as compared to the contralateral control.…”